Cytokinins are a class of N6-substituted adenine derivatives that affect the synthesis of proteins and nucleic acids. In plants, they function as potent hormones that regulate growth and morphogenesis; as constituents of tRNA, they enhance translational efficiency of these molecules; and their biosynthetic pathway is shared by a host of other important products including sterols and steroid hormones. The goal of this project is to set up an experimental system that will facilitate a better understanding of the molecular basis of cytokinin function. Studying cytokinin effects on and synthesis in yeast offers major advantages over analogous experiments in tissue and cell cultures derived from higher plants and animals. The ease of biochemical and especially genetic manipulation, and large-scale culturing; the availability of several relevant mutants, including isopentenyladenosine (i6A) deficient, sterol requiring, and DNA systhesis mutants; and the possibility of utilizing this unicellular eukaryote as a recombinant DNA host make Saccharomyces cerevisiae a powerful tool for the further investigation of this problem. The relationship between i6A synthesis and sterol synthesis will be studied genetically using a number of existing yeast mutants, and biochemically using agents known to inhibit the pathway. The effect of i6A on nucleic acid synthesis will be determined in vitro. The use of temperature sensitive DNA synthetis mutants will help determine the exact nature of any inhibition. The relative contributions to intracellular i6A pools via de novo and tRNA routes will also be assessed. Finally, yeast mutants can be used as recombinant DNA hosts to clone genes in this pathway from yeast and other organisms, affording greater opportunities to study gene function. The significance of this research to health related problems is dyadic. First, the observation that i6A is chemotherapeutic in the treatment of human cancers justifies seeking answers to the question of how it functions. More knowledge of effects on DNA synthesis may shed some light on how this agent induces tumor remission. Secondly, because of the intimate relationship between cytokinin and sterol biosynthesis, dictated by the sharing of several metabolic intermediates, cytokinins can have considerable influence on the production of cholesterol in mammals, including man.